ABSTRACT Fungal species are among the most common causes of nosocomial bloodstream and invasive infections in immunocompromised hosts. Mortality from candidemia can be very high, ranging from 20-40%. Cryptococcus affects approximately 2-3% of solid organ transplant patients and Cryptococcus gattii has emerged as a new cause of pneumonia and disseminated fungal disease. The global emergence of virulent multidrug resistant species such as Candida auris, with a mortality reported to be as high as 68% is particularly worrisome. Culture- based tests to detect Candida infections are slow, relatively insensitive, and may have difficulty distinguishing between C. auris and other related Candida species. Lateral flow antigen tests for Cryptococcus have high sensitivity and specificity but Cryptococcal antigen detection is available only as a standalone assay that is not part of a larger fungal panel; thus, Cryptococcus infections may be missed in the absence of clinical suspicion. Nucleic acid amplification based Candida assays do not have the sensitivity required to detect a Candida infection directly from a patient blood sample (without pre-culture amplification) except for the T2Candida test (based on magnetic resonance technology), which is prohibitively expensive, slow, low throughput and is only able to distinguish among a limited number of Candida species. A more widely available, more rapid and user friendly, and less expensive test for fungal blood stream infections has the potential to greatly impact patient care and improve infection control. This proposal will take advantage of years of successful collaborations between the Alland laboratory and Cepheid developing point of care diagnostic assays, including new approaches to large blood volume sample processing and ultra-sensitive DNA sequence detection. We will leverage this experience to create a fungal detection assay with unparalleled performance. Our specific aims are: 1) Assay development. 2) Improved sample processing to maximize test blood volume and enhance assay sensitivity. 3) Development and testing using dried reagent beads. 4) Perform analytic and pre-clinical studies in preparation for large-scale clinical trials.